Methods for displaying polypeptides and uses thereof

Abstract

Provided herein are methods and compositions for displaying a polypeptide on a tubular structure and uses of such displayed polypeptides in the production of antibodies or vaccines.

1 . A method for displaying a polypeptide, the method comprising culturing a bacterial cell that expresses a first intracellular tubular protein, and that comprises a construct encoding a polypeptide of interest fused to a second intracellular tubular protein, under conditions that permit the expression of said first and second intracellular tubular proteins, wherein such expression permits the formation of an intracellular tubular structure, said tubular structure comprising said first and second tubular proteins and displaying a plurality of molecules of said polypeptide of interest. 
     
     
         2 . The method of  claim 1 , wherein the first or second intracellular tubular protein is a VipA polypeptide. 
     
     
         3 . The method of  claim 1 , wherein the first or second intracellular tubular protein is a VipB polypeptide. 
     
     
         4 . The method of  claim 1 , wherein the displayed polypeptide has a conformation substantially similar to the native conformational folding pattern of the protein of interest. 
     
     
         5 . The method of  claim 1 , wherein the displayed polypeptide is more immunogenic than the protein of interest when not displayed in this manner. 
     
     
         6 . The method of  claim 1 , wherein the bacterial cell is from the species  Vibrio  or  Pseudomonas.    
     
     
         7 . The method of  claim 6 , wherein the bacterial cell is  Vibrio cholera  or  Psuedomonas aeruginosa.    
     
     
         8 . The method of  claim 1 , wherein the construct encoding a polypeptide of interest fused to a second intracellular tubular protein is expressed in the bacteria from which the first and second intracellular tubular proteins were derived. 
     
     
         9 . The method of  claim 8 , wherein expression in the bacteria occurs under conditions in which the secretion of Type VI secretion system substrate proteins is detected. 
     
     
         10 . The method of  claim 9 , wherein the Type VI secretion system substrate proteins are Hcp, VgrG and/or orthologs thereof. 
     
     
         11 . The method of  claim 1 , wherein the polypeptide of interest comprises a viral protein, a bacterial protein, a fungal protein or a tumor protein. 
     
     
         12 . The method of  claim 1 , wherein the tubular structure displays a plurality of molecules of the polypeptide of interest as a regular, repetitive structure. 
     
     
         13 . A method of vaccinating a subject, the method comprising:
 (a) isolating a tubular structure from a cultured bacterial cell that expresses a first intracellular tubular protein, and that comprises a construct encoding a polypeptide of interest fused to a second intracellular tubular protein,   wherein the bacteria cell is cultured under conditions that permit the expression of said first and second intracellular tubular proteins, wherein such expression permits the formation of an intracellular tubular structure, said tubular structure comprising said first and second tubular proteins and displaying a plurality of molecules of said polypeptide of interest, and   (b) administering the isolated tubular structure of step (a) to a subject, thereby vaccinating a subject against the displayed polypeptide of interest.   
     
     
         14 . The method of  claim 13 , wherein the first or second intracellular tubular protein is a VipA polypeptide. 
     
     
         15 . The method of  claim 13 , wherein the first or second intracellular tubular protein is a VipB polypeptide. 
     
     
         16 . A kit for displaying a polypeptide of interest, the kit comprising:
 a vector encoding a first intracellular tubular protein of interest, the vector comprising a multiple cloning site for cloning a polypeptide of interest such that upon expression in a cell, a fusion protein comprising the first intracellular tubular protein of interest and the polypeptide of interest is expressed, and instructions therefor.   
     
     
         17 . The kit of  claim 16 , further comprising a bacterial cell expressing a second intracellular tubular protein that when expressed with said first intracellular protein forms an intracellular tubular structure comprising said first and second intracellular tubular proteins that displays said polypeptide of interest.